Ion channels in mammalian systems are the subject of intensive scientific investigation because of the importance and the variety of their functions.1 Such channels are located in the phospholipid bilayer membranes of cells and permit the transport at a high rate of a particular cation or anion. Natural ion channels are generally understood to be polypeptide or protein structures. These large molecules form pores within membranes that provide diffusion pathways for ionic species that are exquisitely controlled and specific. These diffusion pathways control the movement of ion currents in membranes generating membrane potentials and currents that influence all aspects of cellular physiology. Channel proteins have “open” and “closed” states that are sensitive to membrane potential (voltage gating) or to the presence or absence of regulatory molecules (ligand gating).2 The channel molecules typically exhibit selectivity for one particular cation or anion, in vivo. Often, they can permit the selected ion to be transported preferentially in one direction (rectification). In combination, these characteristics allow ion channels to provide the rapid and integrated control of cellular activity required for mammalian function.
The functions of ion channel proteins have generally been studied by using proteins reconstituted from natural sources or expressed by cloning techniques. The proteins thus obtained are then studied for ion conduction in a lipid bilayer membrane. The lipid bilayer prevents ion diffusion except through the channel and allows detailed study of the currents that result. In this configuration, picoAmp currents can be studied allowing characterization of single channel molecules. Using these methods, it is possible to determine channel conductance, ion selectivity, gating kinetics and to study modulation of channel activity. Using these techniques, the characteristics of many different ion channels have been determined showing a diversity of properties that includes cation and anion specific channels for both monovalent and polyvalent ions.
A common failing of existing synthetic channels is their inability to provide the degree of mimetic fidelity to native channels that would permit meaningful use of the synthesized versions in biomedical applications.3-5 The peptide ion channels produced by the synthetic approach of Lear et al.4 were not intended to be mimetic, but were simply designed to show ion transport. These compounds were formed from repeating units involving only leucine and serine. Mutter et al. have suggested that synthetic channel peptides may be made by linking amphipathic α-helix and β-sheet peptides to a synthetic peptide template.6 The compounds of Mutter et al. were less selective and efficient than their natural channel counterparts.
During the past two decades, numerous attempts have been made to develop synthetic ion channels. These include the total synthesis of a simple but natural channel,7 non-naturally occurring peptide aggregates,8 cyclic peptide nanotubes,9 hybrid crown ether-peptide structures,10 oligomeric structures,11 and completely synthetic compounds called “hydraphiles.”12 Other synthetic channel structures that have been assessed for their ability to transport cations may be identified from recent reviews on the subject.13 Fewer attempts have been reported to develop anion channels.14 
The discovery of functional synthetic ion channels would provide a means to investigate and modulate the regulation of membrane potentials, intracellular pH, cell volume, signal transduction, and transepithelial ion transport in tissues containing absorptive or secretory epithelia under normal and disease conditions. Discovery of novel synthetic ion channel compounds satisfies a need in the art by providing new compositions useful in conditions and diseases wherein transport of ions plays a role.
Such channel assemblies can be used, e.g., in the treatment of diseases such as cystic fibrosis (CF) and adult polycystic kidney disease (APKD). Cystic Fibrosis (CF) is the most common fatal genetic disease in Caucasians.15 Approximately one in every 2,500 Americans of European descent is born with the disease. Despite current standard therapy, the median age of survival is only 26 years. Disease of the pulmonary airways is the major cause of morbidity and is responsible for 95% of the mortality.
Based on both genetic and molecular analyses, a gene associated with CF was isolated as part of 21 individual cDNA clones and its protein product predicted.16-18 The protein product of the CF associated gene is called the cystic fibrosis transmembrane conductance regulator (CFTR).17 CFTR is a protein of approximately 1480 amino acids made up of two repeated elements, each comprising six transmembrane segments and a nucleotide binding domain. The two repeats are separated by a large, polar, so-called R-domain containing nucleotide binding sites and multiple potential phosphorylation sites. Based on its predicted domain structure, CFTR is a member of a class of related proteins that includes the multi drug resistance (MDR) or P-glycoprotein, bovine adenyl cyclase, the yeast STE6 protein and several bacterial amino acid transport proteins.17,19 Proteins in this group, characteristically, are involved in the transport of molecules into or out of cells.
CFTR has been postulated to regulate the outward flow of anions from epithelial cells in response to phosphorylation by cyclic AMP-dependent protein kinase or protein kinase C.17,20-24 Hence, for CF and other conditions, the development of synthetic anion channels has significant utility in terms of investigating cellular anion imbalances and in modulating such conditions.
Intracellular compartments are critical to the organization of eukaryotic cells. The vast majority of these compartments are used in the intracellular transport of macromolecules to locations within the cell or in the uptake and processing of extracellular objects. These compartments include the endocytic vesicles, secretory vesicles, phagosomal vesicles and lysosomes primarily but also include a number of associated specialized vesicles. These vesicles have in common the ability to acidify and then employ this acidic pH in their required functions.25 The acidification of these vesicles critically depends upon the chloride permeability of their limiting bilayer membrane. Because they insert into bilayer membranes at low concentrations, in one embodiment, applicants' chloride channel assemblies can be used to study the precise role of chloride permeability in the functions of these vesicles. In addition, the modification of these functions will have key influences upon the secretion, uptake and processing of proteins, bacteria and viruses by eukaryotic cells. Specialized cells such as osteoclasts and renal tubular cells use chloride dependent acidification for bone metabolism and critical detoxification activities.